Lung cancer is the most common cause of cancer-related death in both men and women in the United States. Eighty percent of lung cancers are non-small cell lung cancers (NSCLCs) and 30% of NSCLCs are adenocarcinomas, which include a rising population of cancer cases that occur in nonsmokers. Mutations in the Kras oncogene, which signals through a cascade of kinases to promote cellular proliferation, have been identified in 20-30% of NSCLCs. Therapeutic targeting of Kras-driven tumors necessitates the identification of signaling pathways required for oncogenic Kras-driven proliferation. Kras-driven lung cancer cells display higher levels of reactive oxygen species (ROS) than noncancerous lung cells. ROS have been proposed to serve as signaling molecules to activate numerous signaling pathways, including PI3K, ERK1/2 MAPK, and the transcription factors hypoxia inducible factors (HIFs), which promote tumor cell proliferation, angiogenesis, and metastasis. The major form of ROS that participates in signaling in the cytosol is hydrogen peroxide (H2O2), which is generated by its conversion from superoxide (O2-) by copper-zinc superoxide dismutase (SOD1) in the cytosol. We have reported that O2- from mitochondrial complex III and its conversion to H2O2 in the cytosol are required to initiate Kras-induced cellular proliferation and hypoxic activation of HIFs in tumor cells. Presently, it is not known whether complex III-generated O2- or H2O2-dependent signaling in the cytosol is required for oncogenic Kras-driven lung tumorigenicity in vivo. Recent studies indicate that glutamine is a major fuel for the TCA cycle to generate NADH and FADH2. These reducing equivalents donate electrons to the electron transport chain resulting in complex III generated superoxide. We recently reported that Kras-driven tumor cells also utilize glutamine to fuel the TCA cycle. Glutamine can be converted by glutaminase (GLS) to glutamate, which enters the TCA cycle through conversion into alpha-ketoglutarate by aminotransferases (GPT2 or GOT2) or glutamate dehydrogenase (GDH). Preliminary data indicate that preventing glutamine entry into the TCA cycle using inhibitors of aminotransferases or RNAi of GPT2 reduces anchorage-independent growth of oncogenic Kras-driven tumor cells. However, it is not known if inhibition of GPT2 would attenuate lung adenocarcinoma in vivo or if GPT2 is dispensable for normal tissues in the adult mouse. The major goal of this grant is to genetically determine whether diminishing complex III generated superoxide, production of cytosolic hydrogen peroxide, and glutamine utilization by the TCA cycle will attenuate tumorigenesis in the oncogenic Kras-driven mouse model of lung adenocarcinoma and in an orthotopic mouse model using human A549 lung adenocarcinoma cells harboring a Kras mutation. PUBLIC HEALTH RELEVANCE: Lung cancer is the most common cause of cancer-related death in both men and women in the United States. The major goal is to determine whether preventing mitochondrial metabolism or production of hydrogen peroxide would be effective in diminishing lung cancer. Positive results would provide a rationale for therapies targeting metabolic enzymes that regulate mitochondrial metabolism.